This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The answers to many questions in biology require knowledge of structures that are undergoing changes between states on time scales not accessible by the powerful structural biology methods of x-ray diffraction and NMR. Kinetic spectroscopy approaches tell us about the rates at which populations interchange and hence about mechanisms. However the range of equilibrium configurations and the structures of the various intermediate states cannot always be determined by these standard methods. In our work we propose new methods to determine structural aspects of these populations and their distributions regardless of how fleeting they are. Such measurements are expected to provide answers to questions regarding the evolution of segments of structures that are not available from other techniques. This is the significance of this work. Vibrational spectroscopy has provided important experimental access to the microscopic aspects of hydrogen bond dynamics in complex systems. The dynamics of O[unreadable]H or O[unreadable]D stretching vibrational modes in water or alcohol oligomers, the vibrations of molecular and atomic aqueous ions, and the N[unreadable]H and C=O stretching modes including those in peptides or proteins in water are very sensitive to and correlated with the structural and dynamical properties of hydrogen bonds. In principle, the shape of the conventional IR absorption spectrum provides information on the equilibrium dynamics of a hydrogen bonded system. However, in many cases the line shapes are determined by population lifetimes and spectral diffusion processes that often cannot be reduced to the unique set of parameters needed to describe the frequencies and amplititudes of coupled solvent nuclear motions. With the help of multidimensional nonlinear spectroscopic techniques in the IR, it has become possible to probe these hydrogen bond dynamics and extract more details on the structures and dynamics with high time resolution. Dynamical information on the O[unreadable]H, O[unreadable]D, and N[unreadable]H stretching modes of intermolecular hydrogen bonded systems has been obtained in the form of vibrational lifetimes, energy transfer, hydrogen bond breaking and reforming rates, and the time dependence of spectral diffusion. The motions of hydrogen bonds in peptides are of importance in biological processes. The amide carbonyl group is very often involved in dynamic hydrogen bonding either to water or to N-H groups or to both. Much remains to be learned from experiments on the vibrational populations and coherences about the dynamics of these hydrogen bonds for a wide range of environments. These chemical exchanges can be directly studied by 2D IR experiments that have wide applicability for the study of such ultrafast dynamics. Furthermore, for a full interpretation of 2D IR this knowledge of the dynamics is necessary. Major objectives of this core project are: - Methods of characterization by 2D IR of the equilibrium and nonequilibrium dynamics and the spatial variations of solvent structure and dynamics. - Development of methods of 2D IR temperature jump experiments applied to continuous structure evolution such as occurs in downhill protein folding reaction. - Methods of optical pump - IR and 2D IR probe spectroscopy to investigate the structures of reaction intermediates of proteins undergoing unfolding transitions. - Hydrogen bond exchange 2D IR methods and the interpretation of 2D IR line shapes including exchanges between vibrationally distinct groups of molecules. - Development of 2D IR methods to measure the frequency correlation functions of multi-amide unit peptides combined with molecular dynamics simulations, ab initio computations and structure/frequency maps. - Methods of dual frequency dynamics and determination of joint correlation functions of C-H, N-H, amide-II and amide-I modes. - Development of direct experimental approaches to expose solvation by dual frequency 2D IR experiments that excite the peptide backbone modes and probe the solvent motions that respond. - Generation of reliable approaches to the measurement and interpretation of vibrational energy transport along peptide backbones.